Cardiovascular disease resulting from atherosclerosis is responsible for over half of the deaths in industrialized nations. Lipid abnormalities associated with high serum levels of total cholesterol, LDL cholesterol, and triglyceride greatly increases the likelihood of development of the clinical manifestations of this disease. Examining the novel mechanisms regulating sterol biosynthesis will yield new information on how cells maintain proper sterol homeostasis. Saccharomyces cerevisiae is a model system for the study of sterol regulation. We have discovered a novel transcriptional regulation of sterol gene expression in yeast. We have found that the transcription of the ERG25, ERG26, and ERG27 genes required for the biosynthesis of zymosterol are up regulated in response to sterol accumulation. The Specific Aims of this application are as follows. (1) Determine whether ERG25, ERG26, and ERG27 are coordinately transcriptionally regulated. We will determine whether ERG25, ERG26, and ERG27 are coordinately regulated in response to various blocks in sterol biosynthesis. We will also determine the mechanism regulating the transcription of these genes with respect to the end product, ergosterol. (2) Determine the promoter sequences required for the sterol-mediated transcriptional regulation of ERG26 gene expression. We will determine the minimal ERG26 promoter sequences required for ERG26 gene expression regulation using ERG26 promoter-lacZ fusion technology. (3) Determine the transcription factors required for the sterol-mediated transcriptional regulation of ERG26 gene expression. We will elucidate the transcription factors required for ERG26 regulation using yeast mutants harboring loss-of-function mutations in known transcription factors and the yeast one-hybrid assay. (4) Determine the role of the yeast oxysterol- binding proteins in the transcriptional regulation of ERG26. Loss-of-function oxysterol-binding protein mutants will be examined for defects in the transcriptional regulation of ERG26. Mutants defective in the yeast Niemann-Pick type C gene will also be examined. (5) Initiate microarray studies that will examine the mechanisms regulating global lipid gene expression. The results of these studies will increase our understanding of the transcriptional mechanisms regulating sterol gene expression in eukaryotes. Moreover, these results may prove useful in the development of new targets for chemotherapeutic intervention with regards to heart disease and Niemann-Pick disease type C.